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Hin-mediated DNA knotting and recombining promote replicon dysfunction and mutation.

Deibler RW, Mann JK, Sumners de WL, Zechiedrich L - BMC Mol. Biol. (2007)

Bottom Line: Here we analyze the effects of recombined and knotted plasmids in E. coli using the Hin site-specific recombination system.We show that Hin-mediated DNA knotting and recombination (i) promote replicon loss by blocking DNA replication; (ii) block gene transcription; and (iii) cause genetic rearrangements at a rate three to four orders of magnitude higher than the rate for an unknotted, unrecombined plasmid.These results show that DNA reactivity leading to recombined and knotted DNA is potentially toxic and may help drive genetic evolution.

View Article: PubMed Central - HTML - PubMed

Affiliation: Interdepartmental Program in Cell and Molecular Biology, Baylor College of Medicine, Houston, Texas 77030-3411, USA. richard_deibler@hms.harvard.edu <richard_deibler@hms.harvard.edu>

ABSTRACT

Background: The genetic code imposes a dilemma for cells. The DNA must be long enough to encode for the complexity of an organism, yet thin and flexible enough to fit within the cell. The combination of these properties greatly favors DNA collisions, which can knot and drive recombination of the DNA. Despite the well-accepted propensity of cellular DNA to collide and react with itself, it has not been established what the physiological consequences are.

Results: Here we analyze the effects of recombined and knotted plasmids in E. coli using the Hin site-specific recombination system. We show that Hin-mediated DNA knotting and recombination (i) promote replicon loss by blocking DNA replication; (ii) block gene transcription; and (iii) cause genetic rearrangements at a rate three to four orders of magnitude higher than the rate for an unknotted, unrecombined plasmid.

Conclusion: These results show that DNA reactivity leading to recombined and knotted DNA is potentially toxic and may help drive genetic evolution.

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Related in: MedlinePlus

Potential models for the Hin-mediated effect. Plasmid pKNOT is recombined by Hin to knot the DNA (a single line represents the central axis of the double helix). In the roadblock model, the knot (or possibly Hin bound to or cleaving DNA) is impassable and stalls polymerase. Alternatively, in the breakage model, knots may break DNA as a result of forces on the plasmid.
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Figure 4: Potential models for the Hin-mediated effect. Plasmid pKNOT is recombined by Hin to knot the DNA (a single line represents the central axis of the double helix). In the roadblock model, the knot (or possibly Hin bound to or cleaving DNA) is impassable and stalls polymerase. Alternatively, in the breakage model, knots may break DNA as a result of forces on the plasmid.

Mentions: We propose two models to explain how Hin blocks the function of the bla gene (Figure 4). These possibilities are not mutually exclusive. The first is the "roadblock" model: Hin, bound to or cleaving DNA, or the knots themselves form an impasse to RNA and/or DNA polymerases. The second is the "breakage" model. Although it is not clear how knots localize in DNA, it has been suggested from numerical studies that knots may spontaneously pull into a tight conformation [44,45]. DNA within the cell is constantly being subjected to a number of pulling forces generated by transcription, replication and segregation. A force (15 pN) comparable to that generated by a single replication or transcription complex [46-49], has been shown to tighten a DNA trefoil to a diameter less than 25 nm [37,50]. Either the roadblock or the breakage model predicts that DNA knots would be mutagenic through replication fork arrest or through a DNA double-strand break. In addition, either could induce the SOS response, which could lead to a genome-wide increase in mutation frequency [51,52].


Hin-mediated DNA knotting and recombining promote replicon dysfunction and mutation.

Deibler RW, Mann JK, Sumners de WL, Zechiedrich L - BMC Mol. Biol. (2007)

Potential models for the Hin-mediated effect. Plasmid pKNOT is recombined by Hin to knot the DNA (a single line represents the central axis of the double helix). In the roadblock model, the knot (or possibly Hin bound to or cleaving DNA) is impassable and stalls polymerase. Alternatively, in the breakage model, knots may break DNA as a result of forces on the plasmid.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC1904230&req=5

Figure 4: Potential models for the Hin-mediated effect. Plasmid pKNOT is recombined by Hin to knot the DNA (a single line represents the central axis of the double helix). In the roadblock model, the knot (or possibly Hin bound to or cleaving DNA) is impassable and stalls polymerase. Alternatively, in the breakage model, knots may break DNA as a result of forces on the plasmid.
Mentions: We propose two models to explain how Hin blocks the function of the bla gene (Figure 4). These possibilities are not mutually exclusive. The first is the "roadblock" model: Hin, bound to or cleaving DNA, or the knots themselves form an impasse to RNA and/or DNA polymerases. The second is the "breakage" model. Although it is not clear how knots localize in DNA, it has been suggested from numerical studies that knots may spontaneously pull into a tight conformation [44,45]. DNA within the cell is constantly being subjected to a number of pulling forces generated by transcription, replication and segregation. A force (15 pN) comparable to that generated by a single replication or transcription complex [46-49], has been shown to tighten a DNA trefoil to a diameter less than 25 nm [37,50]. Either the roadblock or the breakage model predicts that DNA knots would be mutagenic through replication fork arrest or through a DNA double-strand break. In addition, either could induce the SOS response, which could lead to a genome-wide increase in mutation frequency [51,52].

Bottom Line: Here we analyze the effects of recombined and knotted plasmids in E. coli using the Hin site-specific recombination system.We show that Hin-mediated DNA knotting and recombination (i) promote replicon loss by blocking DNA replication; (ii) block gene transcription; and (iii) cause genetic rearrangements at a rate three to four orders of magnitude higher than the rate for an unknotted, unrecombined plasmid.These results show that DNA reactivity leading to recombined and knotted DNA is potentially toxic and may help drive genetic evolution.

View Article: PubMed Central - HTML - PubMed

Affiliation: Interdepartmental Program in Cell and Molecular Biology, Baylor College of Medicine, Houston, Texas 77030-3411, USA. richard_deibler@hms.harvard.edu <richard_deibler@hms.harvard.edu>

ABSTRACT

Background: The genetic code imposes a dilemma for cells. The DNA must be long enough to encode for the complexity of an organism, yet thin and flexible enough to fit within the cell. The combination of these properties greatly favors DNA collisions, which can knot and drive recombination of the DNA. Despite the well-accepted propensity of cellular DNA to collide and react with itself, it has not been established what the physiological consequences are.

Results: Here we analyze the effects of recombined and knotted plasmids in E. coli using the Hin site-specific recombination system. We show that Hin-mediated DNA knotting and recombination (i) promote replicon loss by blocking DNA replication; (ii) block gene transcription; and (iii) cause genetic rearrangements at a rate three to four orders of magnitude higher than the rate for an unknotted, unrecombined plasmid.

Conclusion: These results show that DNA reactivity leading to recombined and knotted DNA is potentially toxic and may help drive genetic evolution.

Show MeSH
Related in: MedlinePlus